DNA damages have been linked to aging, cellular senescence, and development of genetic diseases, including cancer. Cells respond to DNA damage by activating repair and the checkpoint pathways. The checkpoint pathways delay the cell cycle to allow for DNA repair. When the extent of damage is overwhelming, the checkpoint pathways direct the cells to undergo permanent arrest or apoptosis. But the molecular links between the damage recognition, repair and the cell cycle checkpoints, which would be important in determining the fate of a cell, are poorly understood. The checkpoint pathway involves association of the PI3 kinases ATM/ATR with damaged chromatin and subsequent activation of the checkpoint kinases (Chk1 and Chk2) and p53, as well as other regulatory proteins. Despite significant advances, the mechanisms (sensors) that recruit ATM/ATR to damaged-chromatin are not known. Our recent studies revealed that the nucleotide excision repair (NER) protein DDB functions as a sensor in the damage signaling pathway of ATR. The objectives of this proposal are to further investigate the functional link between DDB and ATR. We will test the hypothesis that DDB is a critical partner of ATR in checkpoint activation and genome stabilization. In addition, we will investigate the mechanism by which DDB recruits ATR to the damaged-chromatin and determine whether DDB plays a role in coordinating repair with the checkpoints. DDB is composed to two subunits: DDB1 and DDB2. The DDB2 subunit is mutated in xeroderma pigmentosum (XP-E). We constructed a knockout strain of mice lacking expression of DDB2. These mice recapitulate the key phenotypes of the human disease (XP-E) in that they are highly susceptible to skin carcinogenesis. Moreover, the DDB2-deficient mice develop spontaneous tumors at high frequencies. We will investigate the hypothesis that, in addition to nucleotide excision repair, the tumor suppression function of DDB2 involves the checkpoint pathways related to ATR. The specific aims are: 1. What is the spectrum of spontaneous tumors in DDB2-deficient mice? Are the DDB2 -/- mice deficient in DNA repair and ATR-activated checkpoints? 2. Is DDB required for DNA damage-induced S phase checkpoint? Is DDB required for the genome stabilization function of ATR? 3. How does DDB recruit ATR to damaged-chromatin? Does DDB play any role in coordinating repair with the checkpoints? Does Cop9/signalosome regulate the damage-sensing function of DDB?